1. Field of the Invention
The present invention relates generally to the fields of carbon nanotube chemistry, radioimmunotherapy and other targeted therapies. More specifically, the present invention relates to functionalized single wall nanotube therapeutic compositions, the construction thereof and uses therefor.
2. Description of the Related Art
Single wall carbon nanotubes (SWNT) offer both unusual opportunities as well as challenges. Single wall carbon nanotubes are scalable, while retaining their key properties thus allowing design of a platform suitable for different applications in vivo. In addition, toxicity and clearance are size- and composition-dependent, allowing flexibility of design to reduce possible adverse effects. Single wall carbon nanotubes have enormous aspect ratio, which allows huge amplification of effecter function and altered kinetics of conjugated agents. Additionally, all the carbons atoms, i.e., ˜8000 per 100 nm, are on the surface and therefore available for functionalization allowing attachments of multiple functionalities, in large numbers, simultaneously. This allows simultaneous applications as needed, e.g., for signaling molecules or for therapeutic agents. Single wall carbon nanotubes have a regular and identically repeating structure which should allow construction of regular and repeating functionalization, as well as patterns of functions. Biological processes interacting with the modified single wall carbon nanotubes are likely to recognize the multivalency and repetition as they do for other biological recognition systems.
Single wall carbon nanotubes are inert, stable, flexible, and non-immunogenic. Most larger molecules introduced into living organisms can be recognized by the immune system, and thereafter quickly neutralized upon re-injection, recognized by metabolic systems, and therefore rapidly degraded, or are unstable and denatured within the reducing, warm and physiological environment of the body and tumors. Single wall carbon nanotubes have important electronic properties. The development of biologic sensors, telemetry or crude decision-making devices that could function ex vivo or in vivo may be possible with modified single wall carbon nanotubes.
While there are many potential advantages to the use of single wall carbon nanotubes as a base nanomaterial, there are also possible hurdles and disadvantages. The chemistry necessary to efficiently solubilize single wall carbon nanotubes is beginning to be described but the effects of these identifications on the chemical and electronic properties of the single wall carbon nanotubes are not known.
Although, parent single walled carbon nanotubes, unmodified and in some instances unpurified, have been reported to be toxic, there has been no thorough study concerning the in vivo biological properties of solubilized single wall carbon nanotubes. For example parent single wall carbon nanotubes have been reported to be cytotoxic to human keratinocytes and were also shown to inhibit growth of embryonic rat-brain neuron cells (1). It has been demonstrated separately that parent single wall carbon nanotubes induced the formation of mouse-lung granulomas (2). It has also been reported that single wall carbon nanotubes inhibit the proliferation of human HEK293 cells, induce cell apoptosis and decrease adhesive ability of cells (3). However, for therapeutic and diagnostic applications single wall carbon nanotubes soluble in aqueous media possessing free pendent functionalities for subsequent attachment of drugs or imaging agents are more appropriate. To this effect derivatized single wall carbon nanotubes with pendent peptides or CpG motifs have been shown to cross cell membranes (3). However, at single wall carbon nanotubes concentrations greater than 10 μM cell death was dramatic and no clear mechanism was cited (4). On the other hand, the uptake of solubilized single wall carbon nanotubes (0.05 mg/mL) by endocytosis into a range of cell lines, e.g., HI60, Jurkat, CHO and 3T3 fibroblasts, has been reported and it has been demonstrated that the nanotubes are localized in the endosomes and were non-toxic (5).
Single wall carbon nanotubes are unique among solid-state materials in that every atom is on the surface and hence surface chemistry could therefore be critical to the physical properties of single wall carbon nanotubes and their applications. Single wall carbon nanotubes with aspect ratio approaching 104-105, form a unique class of one-dimensional quantum confined structures exhibiting either semiconducting (sem-) or metallic (met-) behavior are of special interest. Based on their sem- and/or met-character, a number of devices such as field-effect transistors, single electron transistors and computational logic gates have been demonstrated (6).
Moreover, single wall carbon nanotubes have the highest known specific conductivity per unit mass providing the ability to facilitate direct electron-transfer with biomolecules, acting as molecular-scale electrical conduits, and generating unique designing nano-scale biosensors. Similarities between the size scales of enzymes and chemically shortened single wall carbon nanotubes may promote the likelihood of single wall carbon nanotubes to come within electron tunneling distance of enzyme redox sites, improving sensitivity for enzyme labels that generate signals by direct electron exchange and communicate with external data capture devices. Bridging nanotubes with biological systems, however, is a relatively unexplored area, with the exception of a few reports on nanotube probe tips for biological imaging, nonspecific binding (NSB) of proteins, functionalization chemistry for bioimmobilization on nanotube sidewalls, internalization and transport through cell membrane of solubilized single wall carbon nanotubes in in vitro cell cultures and peptide single wall carbon nanotubes constructs for vaccines.
Single wall carbon nanotubes as platforms for investigating surface-protein and protein-protein binding and its subsequent use in cancer immunotherapy has remained completely unexplored. Hence methodologies aimed at the functionalization of nanotubes with specific antibodies and tagging single wall carbon nanotubes with alpha-emitting elements followed by the evaluation of both their biodistribution and specificity in vivo are needed and would have significant therapeutic implications. Early bio-functionalization approaches have relied heavily on modifications of the sidewalls of single wall carbon nanotubes via noncovalent chemistry so as to preserve the sp2 nanotube structure and thus their electronic characteristics (7). The 1,3-dipolar cycloaddition of azomethine ylides with single wall carbon nanotubes results in amine functionalized single wall carbon nanotubes and confers both aqueous solubility and chemical functionalization. These amine functionalized single wall carbon nanotubes are compatible with a myriad of established bioconjugation techniques (8).
There is a significant need in the art for therapeutic methods utilizing functionalized and soluble single wall carbon nanotubes comprising targeting and therapeutic moieties. Specifically, the prior art is deficient in methods of radioimmunotherapy or other immunotherapy or chemotherapy to treat or to prevent a cancer utilizing single wall carbon nanotubes which are functional to target and deliver a therapeutic molecule in vivo. The present invention fulfills this long-standing need and desire in the art.